The dinoflagellate Alexandrium minutum in Cape Town harbour (South Africa): Bloom characteristics, phylogenetic analysis and toxin composition

A novel bloom of Alexandrium minutum occurred in an inner basin of the Cape Town harbour from November 2003 to February 2004. Cellular concentrations reached a maximum of 1.4 × 10⁸ cells l⁻¹ during the mid-December period with corresponding chlorophyll a concentrations of 243 mg m⁻³. Primary productivity measurements conducted during the latter part of the bloom revealed a maximum assimilation number of 11.17 mg C mg Chl a⁻¹ h⁻¹ during the middle of the day. Productivity during this post-peak period was sustained largely by the reduced nitrogen species NH₄ and urea (96%) as measured using ¹⁵N tracer techniques. The large subunit ribosomal DNA sequence of A. minutum isolates from Cape Town harbour was identical to conspecifics collected in Western Europe and in Australia. The composition of tetrahydropurine neurotoxins associated with paralytic shellfish poisoning (PSP) was limited to gonyautoxins (GTX1-GTX4). This profile combined with evidence of a low toxin cell quota (1.5 fmol GTX cell⁻¹) supports a close association of this taxon with other members of the A. minutum species complex, particularly from Europe. Toxin analysis from black mussels collected during this bloom indicated that the accumulated PSP toxins originated from A. minutum and not from Alexandrium catenella as is most often the case along the South African coast.     

Enhancement of growth and gymnodimine production by the marine dinoflagellate, Karenia selliformis

Because of its novel bioactive properties the production of gymnodimine for use as a pharmaceutical precursor has aroused interest. The dinoflagellate, Karenia selliformis produces gymnodimine when grown in bulk culture using GP + selenium medium but the growth rates (μ) and levels of gymnodimine are low (μ, 0.05 days⁻¹; gymnodimine 250 μg L⁻¹ max). We describe the effects of organic acid additions (acetate, glycolate, alanine and glutamate additions and combinations of these) in enhancing growth and gymnodimine production in axenic cultures. The most effective organic acid combinations in decreasing order were: glycolate/alanine > acetate > glycolate. Glycolate/alanine optimised gymnodimine production by prolonging growth (maximum cell yield, 1.76 × 10⁵ cells mL⁻¹; gymnodimine, 1260 μg L⁻¹; growth rate (μ), 0.2 days⁻¹) compared to the control (growth maximum cell yield, 7.8 × 10⁴ cells mL⁻¹; gymnodimine, 780 μg L⁻¹; μ, 0.17 days⁻¹). Acetate enhanced gymnodimine by stimulating growth rate (μ, 0.23 days⁻¹) and the large concentration of gymnodimine per cell (16 pg cell⁻¹ cf. 9.8 pg cell⁻¹ for the control) suggests a role for this compound in gymnodimine biosynthesis. Amending culture media with Mn²⁺ additions resulted in slightly decreased growth in control cultures and increased the gymnodimine while in glycolate/alanine cultures growth was stimulated but gymnodimine production decreased. The results suggest that the organic acid can enhance gymnodimine production by either enhancing growth maximum or the biosynthetic pathway.     

Lack of effect of the nematophagous fungus Duddingtonia flagrans on the development of the dung beetle, Aphodius constans

The nematode-trapping fungus Duddingtonia flagrans may be used as a biological control agent of gastro-intestinal nematode larvae of ruminants by feeding the hosts with fungal spores. This trial was intended to search an eventual detrimental impact of the presence of spores of D. flagrans in high numbers in goat feces on the common dung beetle, Aphodius constans (Coleoptera: Aphodiidae). A. constans eggs were settled in feces derived from grazing goats fed spores at daily dose rates of 0, 0.25 × 10⁶, 0.5 × 10⁶ or 10⁶ spores/kg BW. At the end of the incubation period, the number of adults that have emerged from eggs were counted and compared between dose rates. No difference in emergence rate between treatments can be seen. The presence of D. flagrans spores in goat feces, even in large numbers, did not alter the development of A. constans.     

Alexandrium catenella and Alexandrium minutum blooms in the Mediterranean Sea: Toward the identification of ecological niches

Annual recurrent blooms of the toxic dinoflagellates Alexandrium catenella and Alexandrium minutum were detected from 2000 to 2003 in harbours along the Catalan coast. The interrelation study between the occurrence of the blooms and specific external conditions at the study sites demonstrated that different factors are required for the bloom of each Alexandrium species. Concentrations higher than 10⁵ cells l⁻¹ of A. catenella were only detected in Tarragona harbour. These blooms were associated with water surface temperature between 21 and 25 °C and salinities of around 34 psu or higher than 37 psu. A. minutum appeared widely spread along the Catalan coast, though the most intensive and recurrent blooms of this species were observed in Arenys de Mar harbour. Concentrations of millions of cells per litre of A. minutum were associated with water temperatures below 14 °C and salinities of around 34–36 psu. A. minutum cell densities showed a positive significant correlation with NO₃ but a negative correlation with NH₄. On the other hand, A. catenella blooms dominated when both NO₃ and NH₄ levels were high. The prevailing inorganic nitrogen form (NO₃ vs. NH₄) could explain why these two species rarely coincide in the same harbours. Accumulation of cysts in the sediment was found to be an important potential factor for the recurrence of these species. The 4.3 × 10³ A. catenella cysts cm⁻³ of wet sediment in Tarragona harbour and the 3.02 × 10³ A. minutum cysts cm⁻³ of wet sediment in Vilanova harbour were the highest concentrations observed from the cyst study. Confined waters such as harbours play an important role as reservoirs for the accumulation of cysts and vegetative cells, which contributes to the expansion of these dinoflagellates in the region. However, the particular environmental conditions are also decisive factors of bloom intensity.     

An in vitro Cr release assay for Ascaris suum infective larvae

This report describes the use of ⁵¹Cr release as a measure of the viability of Ascaris suum inefective larvae after treatment with sodium hypochlorite or kalafungin. Inefective larvae of A. suum were treated with varying dilutions of these compounds and percent ⁵¹Cr release and percent visual larval damage were recorded. An 8% solution of sodium hypochlorite released >90% of the ⁵¹Cr and produced >90% visual larval damage. Kalafungin released >32% ⁵¹Cr and produced >60% visual larval damage at a concentration of 1 μg·ml⁻¹. The results demonstrate that both assays are equal indicators of the viability of A. suum infective larvae when the cytotoxic agent is sodium hypochlorite. Kalafungin, an inhibitor of mitochondrial function, is detected more efficiently using percent visual larval damage when compared to ⁵¹Cr release.     

Relationships between larval nutritional experience, larval growth rates, juvenile growth rates, and juvenile feeding rates in the prosobranch gastropod Crepidula fornicata

Planktonic larvae experiencing short periods of starvation or reduced food supply often grow and develop more slowly, have poor survival, fail to metamorphose, metamorphose at smaller sizes, or grow slowly as juveniles. In this study, we examined the impact of short periods of food limitation at various stages of larval development on larval and juvenile growth in Crepidula fornicata. In addition, we considered whether juveniles that were stressed as larvae grew poorly because of reduced rates of food collection due to impaired gill function. For 5 experiments, larvae were either starved for several days beginning within 12 h of hatching or were starved for the same number of days following 1 or more days of feeding at full ration (cells of the naked flagellate Isochrysis galbana, clone T-ISO, at 18×10⁴ cells ml⁻¹). In one experiment, larvae were transferred for 2 or 4 days to seawater with extremely low phytoplankton concentration (1×10⁴ cells ml⁻¹). In all experiments, larvae were returned to full ration following treatment. Control larvae were fed full ration from hatching to metamorphosis. When larvae reached shell lengths of about 900 μm they were induced to metamorphose and then reared individually at full ration in glass bowls, with phytoplankton suspension replenished daily. Larval and juvenile growth rates were determined by measuring changes in shell length (longest dimension) over time. Juvenile feeding rates were determined by monitoring changes in phytoplankton concentration over 2–3 h at the end of the growth rate determinations. In general, larval growth rates for the first 2 days after the resumption of feeding were inversely proportional to the length of time that larvae were starved. However, larval growth rates ultimately recovered to control levels in most treatments. Starving the larvae caused a significant reduction in initial juvenile growth rates (first 3–4 days post-metamorphosis) in most experiments even when larval growth rates had recovered to control levels prior to metamorphosis. Juvenile growth rates were not significantly reduced when larvae were subjected to reduced food availability (1×10⁴ cells ml⁻¹), even for treatments in which larval growth rates were compromised. Mean weight-specific filtration rates for juveniles were significantly reduced (p<0.05) following larval feeding experience in only one or possibly 2 of the 4 experiments conducted. Our data suggest that although larvae of C. fornicata may fully recover from early nutritional stress, the resulting juveniles may exhibit poor initial growth due to impaired gill function, reduced digestive capability, or reduced assimilation efficiency.     

Effect of growth hormone and γ-aminobutyric acid on Brachionus plicatilis (Rotifera) reproduction at low food or high ammonia levels

Growth hormone (GH, 0.0025 and 0.025 I.U. ml⁻¹) and γ-aminobutyric acid (GABA, 50 μg ml⁻¹) enhance rotifer population growth in batch cultures. In order to further understand the mechanism of their actions, we conducted experiments culturing isolated females at low food and high free ammonia levels. At an optimum food level of 7×10⁶ Nannochloropsis oculata cells ml⁻¹ or at low free ammonia level of 2.4 μg ml⁻¹, the F₁ offspring of rotifers treated with GH at 0.0025 I.U. ml⁻¹ had significantly higher population growth rate (r) and net reproduction rate (Ro), and shorter generation time than untreated rotifers. At a lower food level of 7×10⁵ cells ml⁻¹ or at high free ammonia level of 3.1 μg ml⁻¹, rotifers treated with GABA at 50 μg ml⁻¹ had significantly higher r and Ro, and shorter generation time. These results indicate that GABA is effective in enhancing rotifer reproduction when rotifers are cultured under stress whereas GH enhances rotifer reproduction when culture conditions are optimal. Significant effects were also observed in F¹ and F² generations which were not treated with hormones. These data may be useful for treating rotifer mass cultures to mitigate the effects of stress caused by high population densities.     

Feeding on dinoflagellates by intermediate and late stage crab zoeae raised in the laboratory and collected from the field

Hatching stage crab larvae will ingest algae, including non-toxic and toxic dinoflagellates. We determined that later zoeal stages, obtained from both laboratory-raised larvae and natural assemblages, also ingest dinoflagellates and we measured the effects of prey density, prior feeding history and time of exposure to prey on incidence of ingestion. Both stage 1 and later stage larvae exposed to algal prey were examined using epifluorescence for the presence of chl a. Both stage 1 and stage 3 laboratory-raised Cancer oregonensis (Dana) and Hemigrapsus nudus (Dana) ingested both the non-toxic dinoflagellate Prorocentrum micans Ehrenberg and the toxic Alexandrium andersoni Balech, with no difference between the stages. Both species showed higher ingestion of P. micans than A. andersoni. Ingestion of both prey types occurred at prey densities as low as 200 cell ml^− 1 in C. oregonensis and 50 cells ml^− 1 in H. nudus. Samples collected in summer, 2004, provided both stage 1 and late stage Lophopanopeus bellus (Stimpson); stage 1, intermediate, and late stage Fabia subquadrata Dana; and an unidentified porcellanid. Stage 1 L. bellus ingested both prey, while late stage zoeae did not, although the latter apparently were not actively feeding. F. subquadrata fed on both prey, with no difference between early and late larvae. Both stages ingested P. micans more readily than A. andersoni. First evidence of ingestion of P. micans at 600 cells ml^− 1 occurred after only 0.5 h, while it took 2 h for ingestion at 50 cells ml^− 1. The model of larval feeding involving both omnivory and prey discrimination described previously for the hatching stage is sustained throughout zoeal development and is, perhaps, an adaptation to an uncertain prey environment, one that trades opportunism for inefficiency.     

The effects of a harmful alga on bivalve larval lipid stores

Marine invertebrates often have complex life histories that include a swimming planktivorous larval stage, at which time they are vulnerable to a variety of stressors, including those associated with nutritional stress and harmful algal blooms. Lipid stores have been shown to be especially important for post-metamorphic survivorship and growth in a variety of marine invertebrates. We investigated the effects of the harmful brown tide alga Aureococcus anophagefferens on the lipid stores and growth of larvae of the hard clam (northern quahog, Mercenaria mercenaria), a dominant bivalve in many western Atlantic bays and estuaries. M. mercenaria was the dominant bivalve in Great South Bay, Long Island, until the mid-1970s, but very few larvae are presently found in these waters. Recent brown tide blooms have been hypothesized to pose a barrier to recovery of M. mercenaria populations and hinder recent restoration efforts by negatively affecting clam larvae. To test whether a diet of the brown tide alga affects the accumulation of beneficial lipid stores, we fed larvae one of three diets representing equal biovolumes of Isochrysis galbana, a nutritious control alga; A. anophagefferens, the brown tide alga for which nutritional quality is not presently known; or a mixture of the two. Larvae fed only brown tide had significantly less lipid stores than those in the other dietary treatments. In addition, brown tide negatively affected larval size. We also tested for evidence of tradeoffs between larval growth and lipid stores, predicting that when the diet was less nutritious as in the brown tide treatments, larval size and lipids would be negatively correlated. In contrast, we found that larvae fed a mixed algal diet or only A. anophagefferens showed a significant positive correlation between lipid stores and size, suggesting that some larvae were simply better at obtaining food and associated nutrients. Larval success likely depends on a complex interplay between genetic and environmental factors. Our study suggests that poor nutrition associated with a harmful alga can have negative effects on larval size and lipids stores, which in turn are mediated by the inter-individual variability in the ability to grow and accumulate necessary lipid stores. Phytoplankton quality is likely to be important for the sustainability of bivalve populations even when it primarily impacts the larval phase; and a diet of brown tide algae may have lasting legacies for juveniles and adults.     

A remote-query sensor for predictive indication of milk spoilage

Described is application of the remote-query (wireless, passive) magnetoelastic sensor platform for direct detection and monitoring of bacterium contamination of milk within hermetically sealed containers. Specific application is made to the quantification of Staphylococcus aureus ssp. anaerobius (S. aureus) concentrations in milk. S. aureus growth changes milk viscosity, in turn changing the resonance frequency of the liquid immersed sensor allowing S. aureus concentrations of 10³ to 10⁷ cells ml⁻¹ to be directly quantified.     

Effects of Acacia nilotica, A. polyacantha and Leucaena leucocephala leaf meal supplementation on performance of Small East African goats fed native pasture hay basal forages

Optimal utilisation of tannin-rich browse tree fodders including Acacia spp. foliages as crude protein (CP) supplements to ruminants in the tropics is limited by less available information on their feed nutritive potential. Two studies were conducted to: (1) determine rate and extent of ruminal dry matter (DM) degradability (DMD) and (2) investigate effect of sun-dried Acacia nilotica (NLM), A. polyacantha (PLM) and Leucaena leucocephala leaf meal (LLM) supplementation on growth performance of 20 growing (7–9 months old) Small East African male goats (14.6 ± 0.68 kg) fed on native pasture hay (NPH) basal diet for 84 days in a completely randomised design experiment in north-western Tanzania. The goats were randomised into four treatment groups consisting of five animals each. Three supplement diets: 115.3 g NLM (T₂), 125.9 g PLM (T₃) and 124.1 g LLM (T₄), which was used as a positive control, were supplemented at 20% of the expected DM intake (DMI; i.e., 3% body weight) to the three animal groups fed on NPH (basal diet) compared to the animals in a control group that were fed on NPH without browse supplementation (T₁).

NPH had significantly the lowest (P < 0.05) CP of 45.5 g kg⁻¹ DM compared to NLM, PLM and LLM (159, 195 and 187 g kg⁻¹ DM, respectively). NPH had higher (P < 0.05) fibre fractions; lower ruminal DM degradability characteristics and ME than NLM, PLM and LLM. Supplementation of the animals with browse resulted to (P < 0.05) higher average daily weight gains (ADG) of 157.1 g day⁻¹ in T₄ than the animals fed on T₂ (114.3 g day⁻¹) and T₃ (42.9 g day⁻¹), and even to those fed on T₁ (control), which lost weight (−71.4 g day⁻¹). Improved weight gains were mainly due to corrected feed nitrogen (N) or CP due to supplementation of the animals with browse fodder. Too low CP of the NPH would not meet the normal requirements of CP (80 g CP kg⁻¹ DM) for optimal rumen microbial function in ruminants. Higher ADG due to LLM (T₄) and NLM (T₂) supplementation suggest optimised weight gains due to browse supplementation (20% of expected DMI); while lower weight gains from supplementation with PLM (T₃) indicate the possible utilisation of A. polyacantha leaves to overcome weight losses especially during dry seasons.     

Recurrent Pseudo-nitzschia calliantha (Bacillariophyceae) and Alexandrium insuetum (Dinophyceae) winter blooms induced by agricultural runoff

A winter bloom dominated by Pseudo-nitzschia calliantha Lundholm, Moestrup et Hasle (Bacillariophyceae), a potential domoic acid producer, is reported for the first time in the Aegean Sea, Greece, in a semi-enclosed embayment (Kalloni Gulf) surrounded by agricultural land and drained by intermittent rivers. Abundances of this species in the inner part of the Gulf during February were extremely high (max 1.1 × 10⁷ cells l⁻¹). The species Alexandrium insuetum Balech (Dinophyceae) was also found in considerable cell numbers (max 1.4 × 10⁵ cells l⁻¹) during the bloom and reached up to 40% of the total biovolume. This study demonstrates an evident cause and effect relationship between nutrient inflows originating from agricultural activities in the watershed and the development of a potential HAB. The massive bloom formation was observed soon after an episodic rainfall event during the fertilizer application period (December to February). A bloom was also observed the following year, but it was less pronounced due to the fact that rainfalls were more evenly spaced in time and were of moderate intensity.     

Aerobic chromate reduction by chromium-resistant bacteria isolated from serpentine soil

A group of 34 chromium-resistant bacteria were isolated from naturally occurring chromium percolated serpentine soil of Andaman (India). These isolates displayed different degrees of chromate reduction under aerobic conditions. One of the 34 isolates identified as Bacillus sphaericus was tolerant to 800 mg l⁻¹ Cr(VI) and reduced >80% Cr(VI) during growth. In Vogel Bonner broth, B. sphaericus cells (10¹⁰ cells ml⁻¹) reduced 62% of 20 mg l⁻¹ of Cr(VI) in 48 h with concomitant discoloring of yellow medium to white one. Reduction of chromate was pronounced by the addition of glucose and yeast extract as electron donors. In the presence of 4.0 g l⁻¹ of glucose, 20 mg l⁻¹ of Cr(VI) was reduced to 2.45 mg l⁻¹ after 96 h of incubation. Optimum pH and temperature for reduction were 6.0 and 25 °C, respectively. Increase in cell density and initial Cr(VI) concentration increased chromate reduction but was inhibited by metal ions like, Ni²⁺, Co²⁺, Cd²⁺ and Pb²⁺. Experiments with cell-free extracts indicated that the soluble fraction of the cell was responsible for aerobic reduction of Cr(VI) by this organism.     

Allelopathic effects of Ulva lactuca on selected species of harmful bloom-forming microalgae in laboratory cultures

Allelopathic effects of the green macroalgae Ulva lactuca on the growth of three species of red tide microalgae, Heterosigma akashiwo, Alexandrium tamarense, and Skeletonema costatum were tested in laboratory co-cultures precluding the nutrient and light limitation and the effect of high pH. The growth of all three species of microalgae was significantly (p < 0.01) inhibited by fresh U. lactuca. In nutrient replete semicontinuous co-cultures with U. lactuca, H. akashiwo was completely dead in 12 days, and the growth of A. tamarense and S. costatum was reduced by 48 and 46%, respectively by U. lactuca within 12 days. The U. lactuca culture filtrate exhibited a significant (p < 0.05) inhibitory effect on the microalgae in the first 1 or 2 days, but growth resumed in the following days, and S. costatum growth was slightly (p > 0.05) promoted from day 3. The results suggested that the allelopathic compounds are quickly degradable and a long-term inhibition might need the continuous addition of compounds originated from macroalgae. Dried U. lactuca also exhibited inhibitory effects on the microalgae, and the normalized mean growth rates of microalgae decreased with the biomass of dried U. lactuca. The dependent relationships were y = −2.1208x² + 1.0159x + 0.9752 for H. akashiwo, y = 0.7133x² − 3.5813x + 1.1665 for A. tamarense, and y = −0.2114x² − 1.063x + 1.0873 for S. costatum, respectively. The potential feasibility of utilization of dried U. lactuca against red tide microalgae was 2.0 g dry wt L⁻¹. The present study shows that U. lactuca exhibits negative allelopathic effects on harmful bloom-forming microalgae.     

Relative importance of parental and larval nutrition on larval development and metamorphosis of the sea urchin Strongylocentrotus droebachiensis

We examined the relative importance of parental nutritional condition and larval food ration on the rates of development, growth and metamorphosis of larvae of Strongylocentrotus droebachiensis (Müller) in a laboratory experiment. Parents were reared for 22 months on either a high ration of kelp (Laminaria spp., 6 days week⁻¹) supplemented with mussel flesh (Mytilus spp., 1 day week⁻¹) (KM), or a low ration of kelp (1 day week⁻¹) (KL). Larvae were fed either a high ration (5000 cells ml⁻¹) or a low ration (500 cells ml⁻¹) of microalgae (Dunaliella tertiolecta). Larval food ration had a strong effect on the rates of development, growth, and metamorphosis, which were all significantly greater in larvae fed the high ration. Test diameter of settlers also was significantly greater in the high than the low ration. Parental nutritional condition had little or no effect on the rates of development and growth, and no effect on settler size. The rate of metamorphosis was significantly higher in larvae from the KM than the KL treatment in the high but not the low ration (where rates of metamorphosis were similar). Although parental condition generally had a small effect on larval development, our results suggest that when planktonic food is abundant, larvae of adults from nutritionally rich habitats (such as kelp beds) may metamorphose sooner than those of adults from nutritionally poor habitats (such as barrens).     

Human fibroblast culture on a crosslinked dermal porcine collagen matrix

The use of a novel porcine-derived collagen biomaterial as a dermal tissue engineering matrix was examined. The matrix is derived from porcine dermis, and is processed to retain the native collagen (Type 1) and elastin structure. Human primary fibroblasts were cultured on the matrix to examine its potential for creating a dermal replacement. Attachment of fibroblasts on the collagen was compared to tissue culture plastic and PET membranes. Cell proliferation was assessed using the MTT assay and DAPI staining. For seeding densities of 5×10⁴ and 1×10⁵ cells cm⁻², PET and plastic demonstrated >95% attachment of seeded numbers after 3 h. The collagen matrix reached levels >80% after 3–4 h with no influence of the seeding density. Matrix samples with perforating pores of 40 μm diameter were also studied. After 216 h culture in static culture, with media replacement every 3 days, the final cell numbers reached 2.1×10⁵ (perforated) and 2.0×10⁵ cells cm⁻² (unperforated). In comparison fibroblast culture in a perfusion bioreactor, with continuous media replacement, reached 2.3×10⁵ (unperforated) and 2.5×10⁵ cells cm⁻² (perforated) after 216 h.     

Zooplankton feeding ecology: bacterivory by metazoan microzooplankton

Bacterivory by the rotifer Brachionus plicatilis Müller, nauplii and copepodites of the copepods Centropages Krøyer sp. and Acartia tonsa Dana, and the tintinnid Favella panamensis Kofoid & Campbell was examined using fluorescently labelled bacteria (FLB) and epifluorescence microscopy. FLB were < 1 μm in diameter, and were offered at environmental concentrations (1.47−9.08 × 10⁶ cells·ml⁻¹). FLB were visible within rotifers, nauplii, copepodites, and tintinnids, confirming ingestion. Rotifer clearance rates (32–418 μl·animal⁻¹·h⁻¹) exhibited no relation with FLB concentration. In some cases rates of clearance of FLB by rotifers were different with alternative phytoplankton food (Nanochloris Naumann sp.) than in replicates with FLB alone, whereas in other cases presence of alternative food exhibited no clear effects on rates of ingestion of FLB. Clearance rates for all six naupliar stages of A. tonsa nauplii (0–320 μl·animal⁻¹·h⁻¹) were stage-related, with higher rates by NIII-VI nauplii than NI-II nauplii. Nauplii had higher rates of clearance of FLB in the absence of alternative phytoplankton food (Isochrysis Parke sp.). Clearance rates of FLB by a single stage of Centropages sp. nauplii, A. tonsa CI copepodites and F. panamensis (each obtained at only a single food concentration of either 1.5 or 5.0 × 10⁶ cells·ml⁻¹) were within the range of 85–142 μl·animal⁻¹·h⁻¹. These ranges were similar to those of rotifers and A. tonsa nauplii. This is the first report of FLB ingestion by metazoan marine microzooplankton. Although rotifers and ciliates might be expected to ingest small particles such as FLB using ciliary induced feeding currents, the means by which nauplii and copepodites eat FLB is less clear. We propose that they may “eat” bacteria as they “drink” to osmoregulate.     

Critical analysis of application of generalized distance function for optimization of important variables for esterase synthesis by Saccharomyces cerevisiae

The generalized distance function approach is employed to obtain a suitable near optimal conditions of variables. The optimal values of variables (medium constituents, microbiological parameters, and process parameters) have been evaluated separately using single responses (either specific esterase activity or cell mass) as per central-composite-design and multi-responses following generalized distance function approach. The optimal conditions (medium composition (g l⁻¹): dextrose, 13.43; peptone, 7.285; yeast extract, 2.55; and malt extract, 1.695; microbiological parameters: slant age, 39.9 h; inoculum age, 9.6 h; and number of cells, 1.49 × 10⁸ numbers ml⁻¹; process conditions: temperature, 29.9 °C; and pH, 6.2) obtained by generalized distance approach can be considered as a ‘near optimal’ solution of interactive multi-response systems of intracellular esterase synthesis by Saccharomyces cerevisiae.     

Relationship between cyprid energy reserves and metamorphosis in the barnacle Balanus amphitrite Darwin (Cirripedia; Thoracica)

Nauplii batch cultures of Balanus amphitrite were reared with four different diatoms (Skeletonema costatum, Thalassiosira pseudonana, Chaetoceros gracilis, silicate-limited C. gracilis) at three different cells concentrations: 1×10⁵, 5×10⁵, and 1×10⁶ cells ml⁻¹. The cyprid energy reserves were quantified as the ratio of triacylglycerols (TAG) to DNA. Energy reserves of larvae fed on different diatoms at a concentration of 1×10⁶ cells ml⁻¹ were ranked in the order: silicate-limited C. gracilis>C. gracilis>T. pseudonana>S. costatum. There was a significant linear relationship between the TAG content of the diet and cyprid energy reserves. The effect of cyprid energy reserves on metamorphosis to polystyrene surface in the presence and the absence of conspecific settlement factor (SF) was studied after 12, 24, and 48 h of incubation. A strong positive correlation between energy reserves and percent metamorphosis was observed in the absence of SF (r_{12 h}=0.88, r_{24 h}=0.82, r_{48 h}=0.68, P<0.05). A weak positive correlation was observed in the presence of SF (r_{12 h}=0.43, r_{24 h}=0.48, r_{48 h}=0.50, P<0.05). In both treatments, more than 80% of the cyprids with high energy reserves metamorphosed within 24 h. In contrast, a high proportion of cyprids with low energy reserves metamorphosed in response to SF in 24 h. Our results indicate that discriminatory metamorphic behavior of cyprids is closely linked to their TAG/DNA ratio, a proxy for energy reserve.     

Laboratory culture of the Nudibranch Tritonia diomedea bergh (Tritoniidae: Opisthobranchia) and some aspects of its behavioral development

Tritonia diomedea Bergh was reared from oviposition, through metamorphosis to reproductive maturity in the laboratory. The larvae of T. diomedea are planktotrophic and undergo considerable shell growth (from 144.6–329 μ average maximum shell length). Metamorphosis does not require induction, but there may be a preference to metamorphose in the presence of the probable adult prey, a small Virgularia sp. Larvae in cultures fed no food, Dunaliella tertiolecta Butcher Isochrysis galbana Parke, or Monochrysis lutheri Droop did not achieve metamorphic competence at near ambient sea-water temperatures (11.9±1.3 and 13.0±0.8°C). Larvae from cultures fed Monochrysis at room temperature (20.8±1.5°C) or fed a 1 : 1 mixture of Isochrysis and Monochrysis at near ambient sea-water temperatures did metamorphose. Even so, only those larvae fed the 1 : 1 mixture survived more than a few days following metamorphosis. Adult behavioral patterns developed gradually, feeding being first observed at 5 days, swimming in response to NaCl crystals at about 60 days, copulation at about 272 days, and oviposition at about 277 days after metamorphosis. Growth rates were determined for field collected Tritonia diomedea; smaller animals gained and lost weight relatively faster than larger animals.